Building upon evidence that immune manipulation can result in complete and durable regression in select patients with certain metastatic cancers, our efforts have focused on translating these observations to the development of effective immune therapies for hepatocellular cancer (HCC). Essential to this goal is the identification and evaluation of immunotherapeutic targets expressed by HCC. We have begun an active evaluation of putative HCC therapeutic targets including alpha-fetoprotein, members of the cancer-testes antigen family (NY-ESO), and most recently glypican-3 (GPC-3). Our studies have found that GPC-3 mRNA is over-expressed in >90% of fresh HCC tumor specimens and is minimally expressed in normal tissues. Studies evaluating whether this glycoprotein is naturally recognized by T lymphocytes of the human immune system are underway, using traditional techniques involving Tumor Infiltrating Lymphocytes (TIL) derived from surgically resected HCC specimens and also screening of peripheral blood mononuclear cells (PBMC). To facilitate the identification of low frequency and high avidity T cells that recognize novel tumor antigen epitopes, we have pioneered the use of high throughput quantitative gene expression as a tool to rapidly screen for antigen specific T cell reactivity. In pilot studies, we have found this promising technique to be a highly sensitive methodology to identify, stratify, and select melanoma antigen reactive CD8+ T cells from the bulk PBMC of cancer patients. We are applying this method to the screening of potential HCC antigens, and are in the process of adapting the assay as a novel immune monitoring tool for future immunotherapy clinical trials. We have active pre-clinical studies evaluating the role of demethylating agents as a means to modulate the expression of tumor antigens on HCC to enhance their recognition by immune cells. Further pre-clinical efforts are aimed at evaluating the use of non-specific immune stimulating agents, such as, interleukin-2 and anti-CTLA-4 antibody as possible adjunct therapies for the treatment of HCC. With growing evidence suggesting the importance of regulatory T cells in suppressing natural immune responses against tumors, including HCC, we have begun to evaluate their role in HCC and potential therapies that may abate their suppression.